The role of actin in fertilization and embryonic development is an important but poorly understood problem in reproductive biology. Actin has been implicated in a number of developmental events, including fertilization, incorporation of the sperm into the egg, cytokinesis, and morphogenetic movements of cells and subcellular organelles. However, the actual function of actin in these events is not understood. The long-term objective of the proposed research is a more complete understanding of the function of actin in fertilization and early embryogenesis. These studies will examine the role of actin binding proteins in regulating the assembly state and structural organization of actin in the sea urchin egg and embryo. Three specific aims are proposed: 1) Actin binding proteins will be purified from sea urchin eggs by standard techniques, including differential centrifugation, ammonium sulfate precipitation and column chromatography. The interaction of actin binding proteins with actin will be characterized using high and low shear viscometry, sedimentation analysis and electron microscopy. Since both Ca++ and pH have been shown to influence the structural organization of actin in the egg, we will concentrate on actin binding proteins whose activity is Ca++ and/or pH sensitive. 2) Monoclonal and monospecific polyclonal antibodies will be prepared against functional domains on actin binding proteins. 3) These antibodies will be used for the immunolocalization of actin binding proteins in the egg and developing embryo. Actin binding proteins will be localized at the light microscope level by immunoflorescence and at the ultrastructural level using colloidal gold-conjugated antibodies. The results of these studies will provide new information on the role of actin binding proteins in regulating the structural organization and function of actin in the egg and developing embryo. Errors in the control mechanism that regulated the assembly state and structural organization of actin could profoundly alter normal patterns of differentiation and morphogenesis. A better understanding of these control mechanisms will provide new insights into the potential causes of defects in embryonic development.